管傑雄臺灣大學：電機工程學研究所李漢隆Lee, Han-LungHan-LungLee2007-11-262018-07-062007-11-262018-07-062005http://ntur.lib.ntu.edu.tw//handle/246246/53587近幾年來，毛細管電泳(CE)已應用在基因工程上的各種分離技 術、蛋白質的研究和生化科學等領域上。西元2000年七月，美國國家衛生院宣佈完成人體基因計劃(Human Genome Project)的第一階段，主要歸功於毛細管電泳分析法的進步與發展，由於微量且快速等優點，加速了基因定序的提早完成。但傳統的毛細管電泳儀器體積龐大，常限制了毛細管電泳的應用範圍。近來結合MEMS技術及分析科學而成的微小化分析系統(微流體晶片)漸漸受到重視，其具有樣本需求少、設備體積小、價格低廉及分析時效高等特性。本研究主要目的為架設一套共焦螢光顯微術系統，之後便針對一些系統特性進行評估，在解析度的測試方面，如以波長532微米，數值孔徑0.65的情況下，評估本系統之橫向與縱向解析率大約是0.99微米與2.43微米。將此共焦螢光顯微術系統應用在微流體晶片DNA之檢測，選擇針孔大小適度調整其縱向和橫向的解析度，達到檢測分離的DNA片段目的，並探討電動力學中的電滲透流、電泳現象和聚合物分子篩效應對於DNA片段分離的影響。In recent years, CE has been applied in the field of gene engineering and the research of protein and biochemistry. In July, 2000, American Sanitation Bureau announced that they had made the first stage of Human Genome Project. They ascribed the success to the development and progress of CE because the technique had the advantages of saving amount and time. In the past, the instruments of conducting CE were usually big and thus they are not easy to be used in other fields. Recently, the CE chip that integrates MEMS and analysis science is gradually concerned because it has the advantages of requiring fewer samples, having a small volume, a lower price, and a higher efficiency. And the main purpose of this research aims at setting up a Coufocal Fluerescence Microscopy System and then using this system to assess some other particular systems. In the resolution test, under the circumstances that the wavelength is 532 nm and the aperture is 0.65, the trausverse and axial resolution of this system is about 0.99um and 2.43um. We will apply this Coufocal Fluorescence Microscopy System to the DNA test of CE chip and discuss the influence of the electroosmotic and electrophoresis in electronic dynamics and the effect of polymer molecule screen upon the DNA separation.中文摘要…………………………………………………………………..………...Ⅰ 英文摘要…………………………………………………………………………….Ⅱ 目錄………………………………………………………………………………….Ⅲ 表目錄索引………………………………………………………………………….Ⅴ 圖目錄索引………………………………………………………………………….Ⅵ 第一章 緒論 1.1. 研究背景.............………………………………………...……...................1 1.2. 研究動機…..…….....…………………………............................................8 1.3. 研究目的…..……....................…………...................................................10 第二章 實驗原理 2.1. 共焦顯微術原理............……………………….........................................11 2.2. 共焦顯微術之理論模型 2.2.1. 光學成像系統的數學模型…..………..........................................15 2.2.2. 共焦成像系統的橫向與縱向解析率………………………........19 2.2.3. 針孔大小對共焦顯微系統的影響……………………………....24 2.3. 利用共焦螢光顯微術偵測微流道的偵測原理 2.3.1. 縱向偵測原理圖示說明………………………………………...26 2.3.2. 橫向偵測原理圖示說明………………………………………...28 2.4. 螢光的特性 2.4.1 Stokes位移(Stokes shift)………………………………….……..29 2.4.2 螢光的放射光譜……………………………….………………...30 2.5. 毛細管電泳晶片中的電動力學現象 2.5.1 電滲透流………………………...…………………..…………...32 2.5.2 電泳…………………………...………………………..………...33 2.5. 聚合物分子篩效應………………………………………………………..34 第三章 系統架設與校正 3.1. 光學系統與元件……………..…………….……………………...….…..35 3.2. 三軸平移台控制系統.………………..…..................................................45 3.3. 系統特性量測與校正 3.3.1 三軸平移台系統速度量測與校正…..…….………………..…...47 3.3.2 縱向解析度量測結果………………..…….….…………….…...48 3.3.3 橫向解析度量測結果……………..………………..…….……...50 第四章 實驗結果分析與討論 4.1. 實驗樣品製作與準備………………………………...…………………...54 4.2. 毛細管電泳晶片中DNA片段分離狀況分析 4.2.1. 毛細管電泳晶片之螢光偵測結果…….........................................58 4.2.2. 電滲透流對DNA片段分離之影響……….....................................60 4.2.3 電泳對DNA片段分離之影響…………………...……………...62 4.2.4 聚合物分子篩效應對DNA片段分離之影響...……………...…66 第五章 結論................…………………………………........................................70 參考文獻.........................................…………………………………………….……712402533 bytesapplication/pdfen-US共焦螢光顯微術微流體晶片Confocal Fluorescence MicroscopyCE chipthesishttp://ntur.lib.ntu.edu.tw/bitstream/246246/53587/1/ntu-94-R92921044-1.pdf